HLM006474

Cell experiment:

The antiproliferative activity of compounds and their combinations is evaluated using a high-throughput CellTiter-Blue cell viability assay. For the assays, 1000 cells in 24 µL are plated in 384-well plates and incubated overnight at 37°C, 5% CO2. The next day, the drugs are diluted in media and 6 µL of these dilutions added to appropriate wells using an automated pipetting station. Four replicate wells are used for each drug concentration. The cells are incubated with the drug for 120 hours, at which time, 5 µL CellTiter-Blue reagent is added. Cell viability is assessed by the ability of the remaining treated cells to bioreduce resazurin to resorufin (579 nm Ex/584 nm Em). Fluorescence is read with a Synergy HT microplate reader. IC50s are determined using a sigmoidal equilibrium model regression using XLfit version 4.3.2 and is defined as the concentration of drug required for a 50% reduction in growth/viability. For 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) cell viability assays, CellTiter 96 AQueous One Solution is added according to vendor instructions to cells for 2 hours following treatment with drug at the noted dose for 72 hours. All experiments are performed in triplicate and repeated at least three times[2].

References:

[1]. Ma Y, et al. A small-molecule E2F inhibitor blocks growth in a melanoma culture model. Cancer Res. 2008 Aug 1;68(15):6292-9.
[2]. Kurtyka CA, et al. E2F inhibition synergizes with paclitaxel in lung cancer cell lines. PLoS One. 2014 May 15;9(5):e96357.